Cytochrome c oxidase (CcO) is the key, terminal respiratory enzyme responsible for harnessing the oxidative power of dioxygen necessary for energy production in aerobic organisms. Chemically, the net product of the reaction is two equivalents of water, generated by exhaustive reduction of dioxygen concomitantly with pumping of H+ across the mitochondrial membrane to generate a chemiosmotic potential. Due to CcO's essential biochemical function, the pathology associated with dysfunctional mutation generally proves incompatible with life. For example, severe infantile diseases characterized by hypotonia and cardioencephalomyopathy have been linked to attenuated CcO function. Similarly, mutations in CcO caused by the aging process in older populations contribute to reduced muscle mass among other degenerative processes, making study of CcO an engaging target. The active site of dioxygen reduction in CcO is unique among metalloproteins, which attracts the attention of synthetic and biochemists alike with the common goal of understanding the structure-function relationships that ultimately lead to the intrinsic reactivity. Despite decades of intense research effort, direct spectroscopic probing of kinetically trapped enzymatic intermediates has not provided information in a level of detail necessary for characterization of the mechanism of O-O bond rupture. Therefore, the information must come from small molecule model studies and theory in conjunction with the large body of enzymatic data. Model complexes will be used to test the feasibility of a peroxo intermediate during CcO catalysis as predicted by theory. Combined with direct spectroscopic probing of CcO state PM, this study will further test the possibility of an active site tyrosine functioning as a hydrogen atom donor. Finally, the effect of spin state, spin coupling, and copper ion denticity will shed important insight into the direction of the oxygen cleavage coordinate and provide a function based rationale for active site structural elements. The studies require application of advanced spectroscopies such as resonance Raman, X-ray and optical magnetic circular dichroism, electron paramagnetic resonance, and X-ray absorption to be combined with electronic structure methods such as DFT to relate the results to enzymatic catalysis. Thus, the study entails a rich training component for the Trainee. PUBLIC HEALTH RELEVANCE: These studies will yield molecular level details concerning dioxygen reduction by CcO, insight useful for understanding pathogenesis of CcO mutation and perhaps as a basis for "green" chemical catalysts.